Lettuce variety multigreen 50

ABSTRACT

The present invention relates to plants of a lettuce variety NUN 09050 LT (referred to as “MULTIGREEN 50”) and, seeds and progeny thereof. The invention further relates to methods for producing a lettuce plant by traditional breeding methods. Moreover, the invention relates to a method for producing a lettuce plant containing in its genetic material one or more transgenes.

This application claims priority under 35 U.S.C. 119(a)-(d) to CPVOApplication No. 2011/2207, filed by Nunhems B.V. on 14 Sep. 2011, thedisclosure of which is herein incorporated by reference in its entirety.

FIELD OF THE INVENTION

The present invention relates to the field of plant breeding. Inparticular, the invention provides for a new and distinct variety oflettuce designated NUN 09050 LT (or “NUN 09050” or “09050” or“MULTIGREEN 50”).

All cultivated forms of lettuce belong to the highly polymorphic speciesLactuca sativa that is grown for its edible head and leaves. Lactucasativa is in the Cichoreae tribe of the Asteraceae (Compositae) family.Lettuce is related to chicory, sunflower, aster, dandelion, artichokeand chrysanthemum. L. sativa is one of about 300 species in the genusLactuca.

Fresh lettuce is available in the United States year-round although thegreatest supply is from May through October. For planting purposes, thelettuce season is typically divided into three categories, early, midand late, with the coastal areas planting from January to August, andthe desert regions planting from August to December. Fresh lettuce isconsumed nearly exclusively as fresh, raw product and occasionally as acooked vegetable.

The development of commercial lettuce cultivars requires the developmentof lettuce varieties, the crossing of these varieties, and theevaluation of the crosses. Pedigree breeding and recurrent selectionbreeding methods are used to develop cultivars from breedingpopulations. Breeding programs combine desirable traits from two or morevarieties or various broad-based sources into breeding pools from whichcultivars are developed by selfing and selection of desired phenotypes.The new cultivars are crossed with other varieties and the hybrids fromthese crosses are evaluated to determine which have commercialpotential.

Lifestyles change and the demand from restaurants and catering firms forcolorful and interesting garnish for sandwiches and ready-to-useprocessed salads continue to rise. As a result, there is a demand forbreeding companies to develop new varieties with specific shapes ofleaves, specific average size of leaves, prominent color and a widevariety of texture, as well as good yield.

SUMMARY OF THE INVENTION

The invention provides for a new Cutting/Leaf lettuce variety designatedNUN 09050, representative seed of said variety having been depositedunder Accession Number NCIMB ______, and plant parts of the new varietysuch as (harvested) leaves, or parts of the leaves. Non-limitingexamples for parts of said plant are microspore, pollen, ovary, ovule,flower, stalk, leaf, head, shoot, shoot tip, seed, embryo, embryo sac,cutting, root, root tip, pistil, anther, cotyledon, hypocotyl,meristematic cell, stem, cell, in vitro cell cultures or tissuecultures, protoplast, meristem, fruit, petiole, bud or parts of any ofthese such as parts of leaves. In some embodiments, parts of a lettuceplant designated NUN 09050 are provided which are suitable for sexualreproduction or vegetative reproduction, or a tissue culture or cellculture of a lettuce plant designated NUN 09050 is provided.

The invention also provides for seeds of the new variety, representativeseed of said variety having been deposited under Accession Number NCIMB______ or PTA ______, a plant, or a part thereof (such as a lettuce heador leaves), produced by growing said seed.

In a further aspect, the invention provides a plant which is regeneratedfrom a part of the lettuce plant designated NUN 09050 that is suitablefor vegetative reproduction or a plant which is regenerated from a cellor tissue culture of a plant designated NUN 09050.

Also provided are one or more progeny plants (offspring or descendants)of a lettuce plant designated NUN 09050 obtained by further breedingwith said variety designated NUN 09050. Said progeny plant(s) has/haveessentially all physiological and morphological characteristics ofvariety NUN 09050 when grown under the same environmental conditions. Inone embodiment, said progeny plant(s) has/have at least intermediateshape of cotyledons to fourth leaf stage (see USDA criteria); coarselydentate apical margin of fourth leaf (see USDA criteria); a moderateincision depth of margin of mature leaves (see USDA criteria); shallowlydentate indentation of mature leaves (USDA criteria).

Moreover, also an Essentially Derived Variety (EDV) of a lettuce plantdesignated NUN 09050 is provided, e.g., an Essentially Derived Varietyof NUN 09050 having one or two physiological and/or morphologicalcharacteristics which are different from those of NUN 09050 and whichotherwise has essentially all physiological and morphologicalcharacteristics when grown under the same environmental conditions of alettuce plant designated NUN 09050 obtainable by selecting a natural orinduced mutant, or a somaclonal variant from a population of plantsdesignated NUN 09050.

In another aspect the invention refers to packages, e.g., a container, abag and the like, comprising at least one of the following: seeds orseed pellets of lettuce variety designated NUN 09050, lettuce plant(s)designated NUN 09050, parts thereof (e.g. heads or loose leaves),progeny of a lettuce plant designated NUN 09050, parts thereof, EDV of aplant designated NUN 09050 or parts thereof.

Also provided is a food or feed product comprising at least a part of alettuce plant designated NUN 09050, progeny thereof or a EDV thereof.

In further aspects, the invention provides methods of producing alettuce plant, comprising crossing a lettuce plant designated NUN 09050with a second lettuce plant one or more times and/or selfing the lettuceplant designated NUN 09050 one or more times, and selecting progeny fromsaid crossing and/or selfing.

Also provided is a method of producing a lettuce plant derived from NUN09050 comprising the steps of:

-   -   (a) preparing a progeny plant derived from NUN 09050 by crossing        the plant designated NUN 09050 with itself or with a second        lettuce plant;    -   (b) crossing the progeny plant with itself or a second lettuce        plant to produce a seed of a progeny plant of a subsequent        generation;    -   (c) growing a progeny plant of a subsequent generation from said        seed and crossing the progeny plant of a subsequent generation        with itself or a second lettuce plant; and    -   (d) repeating step b) and/or c) for at least 1 more generation        to produce a lettuce plant derived from NUN 09050.

In another aspect the invention provides a method of producing a hybridlettuce seed comprising crossing a first parent lettuce plant with asecond parent lettuce plant and harvesting the resultant hybrid lettuceseed, wherein said first parent lettuce plant or said second parentlettuce plant is a lettuce plant designated NUN 09050.

Also provided is hybrid lettuce seed (from which a hybrid plant can begrown) produced from crossing a first parent lettuce plant with a secondparent lettuce plant and harvesting the resultant hybrid lettuce seed,wherein said first parent lettuce plant or said second parent lettuceplant is a plant designated NUN 09050. Moreover, also provided arehybrid lettuce plants which are grown from a hybrid lettuce seed whereofone parent is a plant designated NUN 09050.

In another aspect the invention provides a method of introducing asingle locus conversion into a plant designated NUN 09050,representative seed of which having been deposited under AccessionNumber NCIMB ______or PTA ______, comprising

-   -   (a) crossing a plant designated NUN 09050 with a second plant        comprising a desired single locus to produce an F₁ progeny        plant;    -   (b) optionally selfing said F₁ progeny plant to produce an F₂        progeny plant having said single locus;    -   (c) crossing said F₁ or F₂ progeny plant of step (a) or step        (b), respectively, with a plant of NUN 09050, representative        seed of which having been deposited under Accession Number NCIMB        ______ or PTA ______, to produce backcross progeny plants;    -   (d) selecting backcross progeny plants that have the single        locus and otherwise essentially all physiological and        morphological characteristics when grown under the same        environmental conditions of a plant designated NUN 09050; and    -   (e) repeating steps (c) and (d) one or more times in succession        to produce selected second or higher backcross progeny plants        that comprise the single locus and otherwise comprise        essentially all physiological and morphological characteristics        when grown under the same environmental conditions of a plant        designated NUN 09050.

In one embodiment, said single locus confers a trait, wherein the traitis pest resistance or disease resistance such as resistance againstNasonovia ribisnigri NR:0 or resistance against Nasonovia ribisnigriNR:1. Further pest or disease resistances are, e.g., resistance againstdowny mildew, e.g., at least one race selected from the group consistingof races B1:1 to 28 and other known Bremia lactucae races, Sclerotiniarot, Botrytis, powdery mildew, anthracnose, bottom rot, corky root rot,lettuce mosaic virus, big vein, lettuce aphid, beet western yellows andaster yellows, Sclerotinia minor (leaf drop), Sclerotinia sclerotiorum(leaf drop), Rhizoctonia solani (bottom drop), Erysiphe cichoracearum(powdery mildew), Fusarium oxysporum f. sp. lactucae (Fusarium wilt),lettuce infectious yellows virus (LIYV), lettuce mosaic virus (LMV),Cucumber mosaic virus (CMV), Beet western yellows virus (BWYV), andAlfalfa mosaic virus (AMV).

In another embodiment the trait conferred by said single locus isanother trait, such as a morphological or physiological trait, herbicideresistance, stress tolerance or stress resistance (e.g. droughttolerance), enhanced yield, etc.

A further aspect of the invention relates to a method of producingtransgenic progeny of a plant designated NUN 09050 wherein said progenyhas at least the essential physiological and morphologicalcharacteristics when grown under the same environmental conditions ofthe variety designated NUN 09050 and further comprises a desired trait,said method comprising transforming a NUN 09050 plant with at least onetransgene that confers said desired trait or crossing a plant designatedNUN 09050 with a lettuce plant comprising said desired trait so that thegenetic material of the progeny that results from the cross contains thetransgene(s). Moreover, also transgenic plants produced by this methodare provided herewith.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 depicts the shapes and colors of fourth leaves of MULTY, NUN09050 and GRAND RAPIDS.

DETAILED DESCRIPTION

All patent and non-patent literatures cited herein are incorporated byreference in their entireties.

“Lettuce” refers herein to plants of the species Lactuca sativa L.

The terms “NUN 09050 LT”, “lettuce plant designated NUN 09050”, “NUN09050”, “MULTIGREEN 50” or “variety designated NUN 09050” refer to alettuce plant/variety of lettuce, representative seed of which havingbeen deposited under Accession Number NCIMB ______ or PTA ______.

“USDA descriptors” are the plant variety descriptors described forlettuce in the “Objective description of Variety Lettuce (Lactuca sativaL.)”, ST-470-1 (dated Jul. 1, 2009) as published by U.S. Department ofAgriculture, Agricultural Marketing Service, Science and Technology,Plant Variety Protection Office, Beltsville, Md. 20705 (available on theworld wide web at www.ams.usda.gov/AMSv1.0/) and which can be downloadedfrom the world wide web atwww.ams.usda.gov/AMSv1.0/getfile?dDocName=stelprdc5069208.

“UPOV descriptors” are the plant variety descriptors described forlettuce in the “Guidelines for the Conduct of Tests for Distinctness,Uniformity and Stability,” TG/013/10 (Geneva 2006), as published by UPOV(International Union for the Protection of New Varieties and Plants,available on the world wide web at upov.int)upov.int/en/publications/tg_rom/tg_index.html. Likewise, “UPOV methods”to determine specific parameters for the characterization of lettuce aredescribed at upov.int.

“RHS” refers to the Royal Horticultural Society of England whichpublishes an official botanical color chart quantitatively identifyingcolors according to a defined numbering system, The chart may bepurchased from Royal Horticulture Society Enterprise Ltd RHS Garden;Wisley, Woking; Surrey GU236QB, UK, e.g., the RHS colour chart: 2007(The Royal Horticultural Society, charity No: 222879, PO Box 313 LondonSW1P2PE; sold by, e.g., TORSO-VERLAG, Obere Grüben 8•D-97877 Wertheim,Article-No.: Art62-00008 EAN-Nr.: 4250193402112).

As used herein, the term “plant” includes the whole plant or any partssuch as plant organs (e.g., harvested or non-harvested leaves, etc.),plant cells, plant protoplasts, plant cell or tissue cultures from whichwhole plants can be regenerated, plant callus, plant cell clumps, planttransplants, seedlings, plant cells that are intact in plants, plantclones or micro propagations, or parts of plants (e.g., harvestedtissues or organs), such as plant cuttings, vegetative propagations,embryos, pollen, ovules, flowers, leaves, heads, seeds, clonallypropagated plants, roots, stems, stalks, root tips, grafts, parts of anyof these and the like, or derivatives thereof, preferably having thesame genetic make-up (or very similar genetic make-up) as the plant fromwhich it is obtained. Also any developmental stage is included, such asseedlings, cuttings prior or after rooting, mature and/or immatureplants or mature and/or immature leaves.

“Cotyledon” refers to one of the first leaves of the embryo of a seedplant; typically one or more in monocotyledons, two in dicotyledons andtwo or more in gymnosperms.

“Tissue Culture” refers to a composition comprising isolated cells ofthe same or a different type or a collection of such cells organizedinto parts of a plant. Tissue culture of various tissues of lettuce andregeneration of plants therefrom is well known and widely published(see, e.g., Teng et al., HortScience. 1992, 27(9): 1030-1032 Teng etal., HortScience. 1993, 28(6): 669-1671, Zhang et al., Journal ofGenetics and Breeding. 1992, 46(3): 287-290).

“Harvested plant material” refers herein to plant parts (e.g., leavesdetached from the whole plant) which have been collected for furtherstorage and/or further use.

“Harvested seeds” refers to seeds harvested from a line or variety,e.g., produced after self-fertilization or cross-fertilization andcollected.

“Head” as used herein refers to lettuce heads, i.e., the plant withoutthe root system, for example substantially all harvested leaves(although this variety is a “loose-leaf” type). Encompassed are immatureleaves (e.g. “baby leaf”) and mature leaves.

The “base” of a plant is the part of a lettuce plant where the leavesare attached to the root system of the plant.

“Core length” of the internal lettuce stem is measured from the base ofthe cut and trimmed head to the tip of the stem.

“Head weight” refers to the mean weight of saleable lettuce head, cutand trimmed to market specifications.

“Head diameter” refers to the mean diameter of the cut and trimmed head,sliced vertically, and measured at the widest point perpendicular to thestem.

“Head height” refers to the mean height of the cut and trimmed head,sliced vertically, and measured from the base of the cut stem to theleaf tip.

“Core Length to Head Diameter Ratio (CLHD Ratio)” refers to the meancore length/head diameter ratio. It is calculated by dividing the meancore length with the mean head diameter. This is an indication of thehead shape and of the ability of a lettuce plant to reduce the amount ofsurface which is on or close to the ground.

“Ground” refers to the surrounding of the aerial tissues of a lettuceplant in which it is grown, or which is placed on the growing medium(e.g. a foil covering the growing medium such as soil but which does notcover the aerial plant, e.g., there is a hole in the foil through whichthe lettuce plant is growing).

A plant having “essentially all the physiological and morphologicalcharacteristics” means a plant having the physiological andmorphological characteristics when grown under the same environmentalconditions of the plant from which it was derived, e.g. the progenitorplant, the parent, the recurrent parent, the plant used for tissue- orcell culture, etc., except for the characteristic(s) derived from aconverted or introduced gene or trait.

As used herein, the term “variety” or “cultivar” means a plant groupingwithin a single botanical taxon of the lowest known rank, whichgrouping, irrespective of whether the conditions for the grant of abreeder's right are fully met, can be defined by the expression of thecharacteristics resulting from a given genotype or combination ofgenotypes, distinguished from any other plant grouping by the expressionof at least one of the said characteristics and considered as a unitwith regard to its suitability for being propagated unchanged.

“Progeny” as used herein refers to plants derived from a plantdesignated NUN 09050. Progeny may be derived by regeneration of cellculture or tissue culture or parts of a plant designated NUN 09050 orselfing of a plant designated NUN 09050 or by producing seeds of a plantdesignated NUN 09050. In further embodiments, progeny may also encompassplants derived from crossing of at least one plant designated NUN 09050with another lettuce plant of the same or another variety or (breeding)line, or wild Lactuca plants, backcrossing, inserting of a locus into aplant or mutation. A progeny is, e.g., a first generation progeny, i.e.the progeny is directly derived from, obtained from, obtainable from orderivable from the parent plant by, e.g., traditional breeding methods(selfing and/or crossing) or regeneration. However, the term “progeny”generally encompasses further generations such as second, third fourth,fifth, sixth, seventh or more generations, i.e., generations of plantswhich are derived from, obtained from, obtainable from or derivable fromthe former generation by, e.g., traditional breeding methods,regeneration or genetic transformation techniques. For example, a secondgeneration progeny can be produced from a first generation progeny byany of the methods mentioned above.

An “Essentially Derived Variety” (EDV) shall be deemed to be essentiallyderived from another variety, “the initial variety”, under the followingcircumstances: (i) it is predominantly derived from the initial variety,or from a variety that is itself predominantly derived from the initialvariety, while retaining the expression of essentially allcharacteristics that result from the genotype or combination ofgenotypes of the initial variety; and (ii) it is clearly distinguishablefrom the initial variety (e.g., one, one or more, two, two or more,three, three or more characteristics are different from the initialvariety); and (iii) except for the differences which result from the actof derivation, it conforms to the initial variety in the expression ofthe essential characteristics that result from the genotype orcombination of genotypes of the initial variety. Thus, an EDV may beobtained for example by the selection of a natural or induced mutant, orof a somaclonal variant, the selection of a variant individual fromplants of the initial variety, backcrossing, or transformation bygenetic engineering. Such a variant may be selected at any time, e.g. inthe field or greenhouse, during breeding, during or after in vitroculture of cells or tissues, during regeneration of plants, etc.

“Plant line” is for example a breeding line which can be used to developone or more varieties.

“Hybrid” refers to the seeds harvested from crossing one plant line orvariety with another plant line or variety.

“F₁ hybrid” refers to the first generation progeny of the cross of twononisogenic plants.

“Crossing” refers to the mating of two parent plants. Equally“Cross-pollination” refers to fertilization by the union of two gametesfrom different plants.

“Backcrossing” refers to a process in which a breeder repeatedly crosseshybrid progeny, for example a first generation hybrid (F₁), back to oneof the parents of the hybrid progeny. Backcrossing can be used tointroduce one or more single locus conversions from one geneticbackground into another. The terms “gene converted” or “conversionplant” in this context refer to plants which are developed bybackcrossing wherein essentially all of the desired morphological andphysiological characteristics of an inbred are recovered in addition tothe one or more genes transferred into the inbred via the backcrossingtechnique or via genetic engineering.

“Regeneration” refers to the development of a plant from cell culture ortissue culture or vegetative propagation.

“Vegetative propagation”, “vegetative reproduction” or “clonalpropagation” are used interchangeably herein and mean the method oftaking part of a plant and allowing that plant part to form at leastroots where plant part is, e.g., defined as or derived from (e.g. bycutting of) leaf, pollen, embryo, cotyledon, hypocotyl, cells,protoplasts, meristematic cell, root, root tip, pistil, anther, flower,shoot tip, shoot, stem, fruit, and petiole. When a whole plant isregenerated by vegetative propagation, it is also referred to as avegetative propagation.

“Selfing” refers to self-pollination of a plant, i.e., the transfer ofpollen from the anther to the stigma of the same plant.

“Single Locus Converted (Conversion) Plant” refers to plants which aredeveloped by plant breeding techniques comprising or consisting ofbackcrossing, wherein essentially all of the desired morphological andphysiological characteristics of a lettuce variety are recovered inaddition to the characteristics of the single locus having beentransferred into the variety via the backcrossing technique and/or bygenetic transformation.

“Transgene” or “chimeric gene” refers to a genetic locus comprising aDNA sequence which has been introduced into the genome of a lettuceplant by transformation. A plant comprising a transgene stablyintegrated into its genome is referred to as “transgenic plant”.

“Substantially equivalent” refers to a characteristic that, whencompared, does not show a statistically significant difference (e.g.,p=0.05) from the mean.

“Average” refers herein to the arithmetic mean.

“First water date” refers to the date the seed first receives adequatemoisture to germinate. This can and often does equal the planting date.

“Maturity date” refers to the stage when the plants are of full size oroptimum weight, in marketable form or shape to be of commercial oreconomic value. This is also the time point when measuring parameters of“mature” leaves.

The term “mean” refers to the arithmetic mean of several measurements.The skilled person understands that the appearance of a plant depends tosome extent on the growing conditions of said plant. Thus, the skilledperson will know typical growing conditions for lettuce of the typesdescribed herein. The mean, if not indicated otherwise within thisapplication, refers to the arithmetic mean of measurements on at least10 different, randomly selected plants of a variety at a requireddeveloping stage (e.g., fourth leaf or mature).

The present invention provides a new type of lettuce (Lactuca sativa)variety, designated NUN 09050. Lettuce variety designated NUN 09050 isof the Cutting/Leaf Type (USDA sheet). The objective of the cross was todevelop a Cutting/Leaf variety with dark green color in mature leaves.

Variety NUN 09050 is most similar to the commercially available varietyMULTY. However, NUN 09050 differs from MULTY in one or more, e.g., atleast two, at least three, at least four, or more, optionally allmorphological and/or physiological characteristics listed in thefollowing (see also Table 1), when grown under the same environmentalconditions:

-   -   when grown under the same environmental conditions, the shape of        cotyledons to fourth leaf stage of NUN 09050 is intermediate        compared to the shape of cotyledons to fourth leaf stage of        MULTY which is spatulate (see USDA criteria);    -   when grown under the same environmental conditions, the apical        margin of fourth leaf of NUN 09050 is coarsely dentate compared        to the apical margin of fourth leaf of MULTY which is        crenate/gnawed (see USDA criteria);    -   color of fourth leaves of NUN 09050 is medium green or darker        whereas the color of fourth leaves of MULTY is light green and        the color of fourth leaves of GRAND RAPIDS is yellow green (see        USDA criteria);    -   when grown under the same environmental conditions the margin of        mature leaves of NUN 09050 has a moderate incision depth whereas        the margin of mature leaves of MULTY has a deep incision depth        (see USDA criteria);    -   when grown under the same environmental conditions, the        indentation of mature leaves of NUN 09050 is shallowly dentate,        whereas the indentation of mature leaves of MULTY is deeply        dentate (see USDA criteria);    -   when grown under the same environmental conditions, the        glossiness of mature leaves of NUN 09050 is moderate whereas        mature leaves of MULTY are more glossy (see USDA criteria);    -   when grown under the same environmental conditions, the head        weight of NUN 09050 at maturity date is at least a factor 1.1 or        even 1.2 or even 1.5 higher than the head weight of MULTY at        maturity date. The date of maturity can vary, for example, the        maturity date in a field trial in Salina (US) was 42 days after        planting for NUN 09050 and 45 days after planting for MULTY;    -   when grown under the same environmental conditions, the core        height from base of head to apex of NUN 09050 is at least a        factor 1.5 higher than the core height from base of head to apex        of MULTY.

The morphological and/or physiological differences between NUN 09050 andother known varieties, such as MULTY or GRAND RAPIDS (which is astandard regional check variety) can easily be established by growingNUN 09050 next to the other varieties (in the same field or greenhouseunder the same environmental conditions), preferably in severallocations which are suitable for lettuce cultivation, and measuringmorphological and/or physiological characteristics of a number of plants(e.g., to calculate an average value (of at least 10 or even more plantswhich were grown under the same conditions) and to determine thevariation range/uniformity within the variety). Comparative varietieswhich can be grown in the same field are GRAND RAPIDS, MULTY and others.These are commercially available varieties.

For example, trials can be carried out in the Netherlands or the USAwhereby e.g., seed characteristics, cotyledon characteristics, fourthleaf characteristics, mature plant characteristics, such as plant headdiameter, head shape, head size, head weight, butt and corecharacteristics, time of bolting (number of days from first water dateto seed stalk emergence), seed stalk characteristics, bolter habit,maturity (earliness of harvest-mature head formation), regional and/orseasonal adaptation, pest and/or disease resistance/susceptibility canbe measured and directly compared. Also resistance against physiologicalstresses, such as tip burn, heat-, drought-, cold-resistance, etc.and/or post-harvest characteristics of heads or leaves can be compared,such as pink rib, russet spotting, rusty brown discoloration, internalrib necrosis (blackheart, grey rib, grey steak) and brown stain can bemeasured using known methods, e.g. as indicated in the USDA descriptors.The morphological and/or physiological characteristics may vary withvariation in the environment (such as temperature, light intensity, daylength, humidity, soil, fertilizer use), which is why a comparison underthe same environmental conditions is preferred. Colors can best bemeasured against The Munsell Book of Color (Munsell Color MacbethDivision of Kollmorgan Instruments Corporation) or using the RHS-Chart(see, e.g., world wide net:http://www.rhs.org.uk/Plants/RHS-Publications/RHS-colour-charts).

Seeds

The present invention does not only provide plants designated NUN 09050.Also provided are seeds of lettuce variety NUN 09050. A representativesample of said seeds (at least 2500 seeds) has been deposited under theBudapest Treaty with Accession Number NCIMB ______ or PTA ______.

Seeds of NUN 09050 are obtainable by, e.g., growing plants from theseeds deposited under Accession number NCIMB ______ or PTA ______ andallowing, e.g., self-pollination and/or cross-pollination and collectingseeds from the resulting plants. The resultant NUN 09050 seeds can begrown to produce plants designated NUN 09050. Moreover, a seed dedicatedNUN 09050 also refers to a seed wherein the plant grown therefrom showsessentially all characteristics of NUN 09050. Seeds of NUN 09050 arealso obtainable by vegetative propagation from plant cells or tissue ofa plant grown from seeds of NUN 09050.

In one embodiment, a plurality of NUN 09050 seeds are packaged intosmall and/or large containers (e.g., bags, cartons, cans, etc.). Theseeds may be pelleted prior to packing (to form pills or pellets) and/ortreated with various compounds, such as seed coatings.

Seed pelleting can be combined with film coating (Halmer, P. 2000.Commercial seed treatment technology. In: Seed technology and itsbiological basis. Eds: Black, M. and Bewley, J. D., pages 257-286).Pelleting creates round or rounded shapes, which are easily sown withmodern sowing machines. A pelleting mixture typically contains seeds andat least glue and filler material. The latter could be, for example,clay, mica, chalk or cellulose. In addition, certain additives can beincluded to improve particular properties of the pellet, e.g., a seedtreatment formulation comprising at least one insecticidal, acaricidal,nematicidal or fungicidal compound can be added directly into thepelleting mixture or in separate layers. A seed treatment formulationcan include one of these types of compounds only, a mixture of two ormore of the same type of compounds or a mixture of one or more of thesame type of compounds with at least one other insecticide, acaricide,nematicide or fungicide.

Formulations especially suitable for the application as a seed treatmentcan be added to the seed in the form of a film coating including alsothe possibility of using the coating in or on a pellet, as well asincluding the seed treatment formulation directly into the pelletmixture. Characteristically, a film coating is a uniform, dust-free,water permeable film, evenly covering the surface of all individualseeds (Halmer, P. 2000. Commercial seed treatment technology. In: Seedtechnology and its biological basis. Eds: Black, M. and Bewley, J. D.,pages 257-286). Besides the formulation, the coating mixture generallyalso contains other ingredients such as water, glue (typically apolymer), filler materials, pigments and certain additives to improveparticular properties of the coating. Several coatings can be combinedon a single seed.

In addition, several combinations with film coating are possible: thefilm coating can be added on the outside of the pellet, in between twolayers of pelleting material, and directly on the seed before thepelleting material is added. Also more than 1 film coating layer can beincorporated in a single pellet. A special type of pelleting isencrusting. This technique uses less filler material, and the result isa ‘mini-pellet’.

Seeds may also be primed. Of all the commercially planted vegetableseeds, lettuce is the most often primed.

Priming is a water-based process that is performed on seeds to increaseuniformity of germination and emergence from the soil, and thus enhancevegetable stand establishment. Priming decreases the time span betweenthe emergence of the first and the last seedlings. Methods how to primelettuce seeds are well known in the art (see, e.g., Hill et alHortScience 42(6): 1436, 2007).

Plant and Parts Thereof.

Also provided are parts of the lettuce plants designated NUN 09050 suchas microspores, pollen, ovaries, flowers, stalks, heads, ovules, leaves,shoots, seeds, embryos, embryo sacs, roots, cuttings, stems, cells,protoplasts, meristems, buds etc. of variety NUN 09050, or parts of anyof these. Such parts may be (part of) sexual reproduction tissues, whichinclude, without limitation microspores, pollen, flowers, ovaries,ovules, embryo sacs and egg cells, or vegetative cells or tissues, whichinclude, without limitation cuttings, roots, stems, cells orprotoplasts, leaves, cotyledons, meristems and buds.

Moreover, there is provided a cell culture or tissue culture of lettucevariety NUN 09050 in which the cell- or tissue culture is derived from atissue such as, for example and without limitation, leaves, pollen,embryos, cotyledon, hypocotyls, meristematic cells, roots, root tips,anthers, flowers, seeds or stems. For example, leaf-, hypocotyl- orstem-cuttings may be used in tissue culture.

Also provided are lettuce plants regenerated from the above-describedplant parts, or regenerated from the above-described cell or tissuecultures, said regenerated plant having essentially all themorphological and physiological characteristics of lettuce variety NUN09050. These plants can also be referred to as “vegetative propagationsof NUN 09050”.

Also provided are harvested leaves and/or heads of NUN 09050 andpackages comprising a plurality of leaves and/or heads of NUN 09050.

Growing Plants

Lettuce plants can be produced by seeding directly in the ground (e.g.,soil such as soil in a field) or by germinating the seeds in controlledenvironment conditions (e.g., greenhouses) and then transplanting theseedlings into the field (see, e.g., Gonai et al., J. of Exp. Bot.,55(394): 111, 2004; Louise Jackson et al, Publication 7215 ISBN978-1-60107-007-4 and Publication 7216 ISBN 978-1-60107-008-1 and theworld wide web at “anrcatalog.ucdavis.edu” search: lettuce forcultivation, harvesting, handling and postharvest methods commonlyused). Lettuce may also be grown in tunnels.

Moreover, lettuce can be grown in hydroponic cultures as described in,e.g., US 2008/0222949.

Hydroponics is the cultivation of plants without soil. There are 6 basictypes of hydroponic systems: Wick, Raft (also called Water Culture), Ebband Flow (also called Flood & Drain), Drip, Nutrient Film Technique, andAeroponic. There are hundreds of variations on these basic types ofsystems, and most hydroponics systems can be described as a variation orcombination of these six types.

In wick systems, plants are grown in a soil-less growing medium and asolution containing water and nutrients is delivered using wicks thatabsorb the solution from a reservoir and deliver the solution to thegrowing medium. The roots of the plants are optionally prevented from orallowed to grow in the solution.

In raft systems, plants are grown in a soil-less growth medium that isfloated by a raft on the surface of a solution containing water andnutrients. The roots of the plants are optionally prevented from orallowed to grow in the solution.

In Ebb and Flow systems, plants are grown in a soil-less growth mediumin a flooding tray. Solution containing water and nutrients isintermittently delivered to the flooding tray and then returned to areservoir. The plant roots are directly or indirectly contacted by thesolution in the flooding tray. Optionally the solution is delivered by apump and returned by gravity.

In drip systems, plants are grown in a soil-less growing medium. Asolution containing water and nutrients is delivered in drips to thegrowing medium. The solution that is not used by the plants is eitherrecycled (recovery systems) or discarded (non-recovery systems). Inrecovery systems, although there often is a reservoir, the plant rootsare typically prevented from growing directly in the solution.

Nutrient film technique (N.F.T.) systems constantly deliver a thin filmof a nutrient and water containing solution. The plants are grown in asoil-less growth medium and the roots are allowed to grow outside themedium into the surrounding air or the plants are grown directlysuspended in the air without a growing medium. The roots that grow inthe air are constantly contacted by the thin film of solution. Typicallythe solution is recycled. Optionally the solution is delivered by a pumpand returned by gravity.

Aeroponic systems deliver the solution as a fine spray. The plants aregrown in a soil-less growth medium and the roots are allowed to growoutside the medium into the surrounding air or the plants are growndirectly suspended in the air without a growing medium. The roots thatgrow in the air are intermittently sprayed or misted with a solutioncontaining water and nutrients. The roots of the plants are optionallyprevented from or allowed to grow in the solution.

Furthermore, different lettuce varieties may be grown as “compositelettuce” (see, e.g., EP 1 197 137 A1).

Plants and Progeny

In another embodiment, plants and parts of lettuce variety designatedNUN 09050 and progeny of lettuce variety designated NUN 09050 areprovided e.g., grown from seeds, produced by sexual or vegetativereproduction, regenerated from the above-described plant parts, orregenerated from cell or tissue culture of the lettuce varietydesignated NUN 09050, in which the reproduced (seed propagated orvegetatively propagated) plant has all or essentially all morphologicaland physiological characteristics of lettuce variety designated NUN09050, e.g., as outlined in table 1. In one embodiment, said progeny ofthe lettuce variety designated NUN 09050 can be modified in one, two,three, four or five characteristics (e.g. different shape of apicalmargin of leaves or further resistances), in which the modification is aresult of, for example and without limitation, mutagenesis ortransformation with a transgene. A progeny can be a first generationprogeny or a progeny up to generation two, three, four, five, six, sevenor even higher.

Furthermore, the invention provides for progeny of variety NUN 09050such as progeny obtained by, e.g., selfing NUN 09050 one or more timesand/or cross-pollinating NUN 09050 with another lettuce plant of adifferent variety or breeding line, or with a lettuce plant of theinvention one or more times. In particular, the invention provides forprogeny that retain essentially all morphological and physiologicalcharacteristics of NUN 09050.

In still another embodiment, there is provided progeny of lettucevariety NUN 09050 produced by sexual or vegetative reproduction, grownfrom seeds, regenerated from the above-described plant parts, orregenerated from the above-described tissue culture of the lettucevariety or a progeny plant thereof, in which the sexually orvegetatively propagated plant has essentially all the morphological andphysiological characteristics of lettuce variety NUN 09050.

Moreover, also an Essentially Derived Variety (EDV) of a lettuce plantdesignated NUN 09050 is provided. In one embodiment, an EDV exhibitsone, two, three or more than three physiological and/or morphologicalcharacteristics which are different from those of NUN 09050 but whichotherwise has essentially all physiological and morphologicalcharacteristics of a lettuce plant designated NUN 09050. In oneembodiment, said EDV has essentially all physiological and/ormorphological characteristics of a lettuce plant designated NUN 09050but has one or two physiological and morphological characteristic(s)which is (are) different from those of the corresponding physiologicaland/or morphological characteristics of a plant designated NUN 09050(i.e. has additional trait(s)).

In one embodiment, NUN 09050 may also be mutated (by e.g. irradiation,chemical mutagenesis, heat treatment, etc.) and mutated seeds or plantsmay be selected in order to change one or more characteristics of NUN09050. Also natural mutants may be identified and used in breeding.Methods such as TILLING and/or EcoTILLING may be applied to lettucepopulations in order to identify mutants. Similarly, NUN 09050 may betransformed and regenerated, whereby one or more chimeric genes areintroduced into the variety. Transformation can be carried out usingstandard methods, such as Agrobacterium tumefaciens mediatedtransformation or biolistics, followed by selection of the transformedcells and regeneration into plants. A desired trait (e.g. genesconferring pest or disease resistance, herbicide, fungicide orinsecticide tolerance, etc.) can be introduced into NUN 09050, orprogeny thereof, by transforming NUN 09050 or progeny thereof with atransgene that confers the desired trait, wherein the transformed plantretains essentially all the morphological and physiologicalcharacteristics of NUN 09050 or the progeny thereof and contains thedesired trait.

The invention also provides for progeny of lettuce variety NUN 09050obtained by further breeding with NUN 09050. In one aspect progeny areF₁ progeny obtained by crossing NUN 09050 with another plant or 51progeny obtained by selfing NUN 09050. Also encompassed are F2 progenyobtained by selfing the F₁ plants. “Further breeding” encompassestraditional breeding (e.g., selfing, crossing, backcrossing), markerassisted breeding, and/or mutation breeding. In one embodiment, theprogeny have all the physiological and morphological characteristics ofvariety NUN 09050 when grown under the same environmental conditions.

In one aspect haploid plants and/or double haploid plants of NUN 09050are encompassed herein. Haploid and double haploid (DH) plants can forexample be produced by anther or microspore culture and regenerationinto a whole plant. For DH production chromosome doubling may be inducedusing known methods, such as colchicine treatment or the like.

The invention also provides a method of producing plants of varietydesignated NUN 09050, or its progeny, or an EDV, or a part thereof,comprising vegetative propagation of a plant designated NUN 09050. Inone embodiment, said vegetative propagation comprises regenerating awhole plant from a part of variety designated NUN 09050. In oneembodiment, said part of a plant is a cutting, root, sterm, cell,protoplast, leaf meristem, bud, cell culture or a tissue culture (e.g.,in vitro meristem culture, see Murakami and Oka, 1996, Plant TissueCulture Letters 13(3): 339). Thus, a vegetative propagated plant (or apart thereof) is provided having at least the essential morphologicaland physiological characteristics of a lettuce plant designated NUN09050 when grown under the same environmental conditions. In someembodiments, said propagated plant has at least one or moremorphological and physiological characteristic in common with a lettuceplant designated NUN 09050. Such characteristics are, e.g. the seedcolor, glossiness, shape of cotyledons etc. as outlined in thisapplication.

The invention also provides for a method of producing a vegetativelypropagated plant of variety designated NUN 09050, or a part thereof,comprising regeneration of said plant from a cell culture or a tissueculture. Also provided are plants which are regenerated from such a cellculture or tissue culture. In one embodiment such plants are haploid ordouble haploid plants of NUN 09050.

In still another aspect, the present invention provides a method ofproducing a plant derived from NUN 09050, the method comprising thesteps of: (a) preparing a progeny plant derived from NUN 09050, whereinsaid preparing comprises crossing a plant of NUN 09050 either as a maleor as a female parent with a second plant, or selfing NUN 09050, orvegetative propagation of NUN 09050, and (b) collecting seeds from saidcrossing or selfing or regenerating a whole plant from the vegetativecell- or tissue culture.

In one embodiment, crossing a plant of NUN 09050 either as a male or asa female parent with a second plant refers to cross-pollination of twoplants of NUN 09050 or of crossing NUN 09050 with an EDV of NUN 09050,or crossing two EDVs of NUN 09050 with each other. In anotherembodiment, crossing a plant of NUN 09050 either as a male or as afemale parent with a second plant refers to cross-pollination of NUN09050 with a plant of another lettuce variety, breeding line or wildLactuca species, e.g. L. virosa or L. serriola.

In yet a further aspect, the invention provides for a method ofproducing a new lettuce plant, e.g., a Fi hybrid. The method comprises,crossing NUN 09050, either as male or as female parent, with a secondlettuce plant which can be a second lettuce plant of the same variety, alettuce plant of a different variety, a breeding line (e.g. an eliteline) or a wild relative of lettuce (e.g., Lactuca virosa or Lactucaserriola) or genetic transformation techniques to produce a progeny ofNUN 09050. The method may further comprise selfing the F₁ to produce anF2 (and optionally further selfing selected F₂ plants to produce an F₃,etc.), and backcrossing an F₁, F₂, F₃, etc. to NUN 09050 or to an EDV ofNUN 09050, to produce a backcross population (BC₁), which may then beselfed to produce a BC₁S₁ population, etc., or may be used to produce aBC₂ population, or other backcross populations.

Thus, in one aspect a method for developing a lettuce plant in a lettucebreeding program is provided, using a lettuce plant of the invention, orits parts, as a source of plant breeding material. Suitable plantbreeding techniques are recurrent selection, backcrossing, pedigreebreeding, line selection, mass selection, mutation breeding and/orgenetic marker enhanced selection. For example, in one aspect, themethod comprises crossing a lettuce plant designated NUN 09050, orprogeny thereof, with a different lettuce plant selected from the groupconsisting of a plant of the same variety, a lettuce plant of adifferent variety, a (breeding) line, or a wild relative of lettuce(e.g., L. virosa or L. serriola), and wherein one or more offspring ofthe crossing are subject to one or more plant breeding techniquesselected from the group consisting of recurrent selection, backcrossing,pedigree breeding, line selection, mass selection, mutation breeding andgenetic marker enhanced selection (see e.g. Allard (1960), John Wiley &Sons, Inc,: Principles of plant breeding, Library of Congress CatalogCard Number: 60-14240).

In still yet another aspect; the present invention provides a method ofproducing a new plant comprising (a) crossing NUN 09050, or a progenyplant thereof, with itself or a second plant to produce a seed of aprogeny plant of a subsequent generation. In further embodiments, themethod may additionally comprise: (c) growing a progeny plant of asubsequent generation from said seed of a progeny plant of a subsequentgeneration and crossing the progeny plant of a subsequent generationwith itself or a second plant; and repeating the steps for an additional3-10 generations to produce a plant derived from NUN 09050. The plantderived from NUN 09050 may be an inbred line, and the aforementionedrepeated crossing steps may be defined as comprising sufficientinbreeding to produce the inbred line. In the method, it may bedesirable to select particular plants resulting from step (c) forcontinued crossing according to steps (b) and (c). By selecting plantshaving one or more desirable traits, a plant derived from NUN 09050 isobtained which possesses some of the desirable traits of the line aswell as potentially other selected traits.

Pedrigee Selection

In one embodiment, Pedrigee selection is used as breeding method fordeveloping a lettuce variety. Pedigree selection, also known as the“Vilmorin system of selection,” is described in, e.g., Allard, 1960,John Wiley & Sons, Inc.: Principles of plant breeding: 119-128, Libraryof Congress Catalog Card Number: 60-14240.

In general, selection is first practiced among F₂ plants. In the nextseason, the most desirable F₃ lines are first identified, then desirableF₃ plants within each line are selected. The following season and in allsubsequent generations of inbreeding, the most desirable families areidentified first, then desirable lines within the selected families arechosen, and finally desirable plants within selected lines are harvestedindividually. A family refers to lines that were derived from plantsselected from the same progeny from the preceding generation.

Using this pedigree method, two parents may be crossed using anemasculated female and a pollen donor (male) to produce F₁ offspring.Lettuce is an obligate self-pollination species, which means that pollenis shed before stigma emergence, assuring 100% self-fertilization.Therefore, in order to optimize crossing, a method of misting may beused to wash the pollen off prior to fertilization to assure crossing orhybridization.

Parental varieties are selected from commercial varieties thatindividually exhibit one or more desired phenotypes. Additionally, anybreeding method involving selection of plants for the desired phenotypecan be used in the method of the present invention.

The F₁ may be self-pollinated to produce a segregating F₂ generation.Individual plants may then be selected which represent the desiredphenotype in each generation (F₃, F₄, F₅, etc.) until the traits arehomozygous or fixed within a breeding population.

Thus, progeny in connection with Pedrigee selection are either thegeneration (seeds) produced from the first cross (F₁) or selfing (S₁),or any further generation produced by crossing and/or selfing (F₂, F₃,F₄, F₅, F₆, F₇, etc.) and/or backcrossing (BC₁, BC₂, BC₃, BC₄, BC₅, BC₆,BC₇, etc.) one or more selected plants of the F₁ and/or S₁ and/or BC₁generation (or plants of any further generation, e.g. the F₂) withanother lettuce plant (and/or with a wild relative of lettuce). Usingcommon breeding methods such as backcrossing or recurrent selection, oneor more specific characteristics may be introduced into NUN 09050, toprovide an EDV of NUN 09050.

In one embodiment, this invention is directed to methods for producing alettuce plant by crossing a first parent lettuce plant with a secondparent lettuce plant wherein either the first or second parent lettuceplant is lettuce NUN 09050. Further, both first and second parentlettuce plants can come from NUN 09050. Still further, this inventionalso is directed to methods for producing a NUN 09050-derived lettuceplant by crossing NUN 09050 with a second lettuce plant and growing theprogeny seed, and repeating the crossing and growing steps with the NUN09050-derived plant from zero to seven times. Thus, any such methodsusing NUN 09050 are part of this invention: selfing, backcrosses, hybridproduction, crosses to populations, and the like. All plants producedusing NUN 09050 as a parent are within the scope of this invention,including plants derived from NUN 09050. Advantageously, NUN 09050 isused in crosses with other, different, lettuce varieties to producefirst generation (F₁) lettuce hybrid seeds and plants with superiorcharacteristics.

It should be understood that the lettuce can, through routinemanipulation of cytoplasmic or other factors, be produced in amale-sterile form. Such embodiments are also contemplated within thescope of the present claims.

General crossing methods for lettuce are, e.g., described in US2009/0271897 A1.

Such methods include but are not limited to:

-   -   Manual removal of anther tubes from flowers, misting the        designated male flowers to wash the pollen off prior to        fertilization and pollen from another variety or donor parent is        then introduced by gently rubbing the stigma and style of the        donor parent to the maternal parent. Tags with the pertinent        information on date and pedigree are then secured to the flowers        in order to keep track. About 3 weeks after pollination, seeds        are harvested when the involucres have matured. The seeds are        eventually sown and in the presence of markers such as leaf        color or leaf margins, the selfed or maternal seedlings or        plants are identified. Generally, in case of crossing with a        plant of a different variety, there are no visible markers and        breeders must wait until the F₂ generations when expected        segregation patterns for the genetic character of interest can        be followed.    -   Use of male sterility systems, such as genetic male sterility        (GMS), see e.g. Hayashi et al. Euphytica Vol 180(3): 429-436) or        cytoplasmic male sterility (CMS). Genetically engineered        sterility is also available. Non-limiting examples of        genetically engineering male sterility in lettuce are by        expression of a ribonuclease gene under the control of a        tapetum-specific promoter (see Reynaerts et al., Scientia        Horticulturae (1993) 55 (1-2): 125-129). Other male sterile        systems include the expression of beta-glucanase via a        tapetum-specific promoter (see Curtis et al., Plant Science        Limerick (1996) 113(1): 113-119).    -   Also provided is a method of producing a hybrid lettuce seed        comprised of crossing a first parent lettuce plant with a second        parent lettuce plant and harvesting the resultant hybrid lettuce        seed, in which the first parent lettuce plant or the second        parent lettuce plant is the lettuce variety NUN 09050.        Accordingly, a hybrid lettuce plant produced from crossing a        first parent lettuce plant with a second parent lettuce plant        and harvesting the resultant hybrid lettuce seed, wherein said        first parent lettuce plant or said second parent lettuce plant        is a lettuce plant designated NUN 09050 is provided, as well.

The invention provides for methods of producing EDVs (EssentiallyDerived Varieties), which retain essentially all morphological andphysiological characteristics of NUN 09050 but which may differ from alettuce plant designated NUN 09050 in one, two, three or more furthermorphological and/or physiological characteristics, but which are stillgenetically closely related to NUN 09050. The relatedness can, forexample be determined by fingerprinting techniques (e.g., making use ofisozyme markers and/or molecular markers such as SNP markers, AFLPmarkers, microsatellites, minisatellites, RAPD markers, RFLP markers andothers). A plant is “closely related” to NUN 09050 if its DNAfingerprint is at least 80%, 90%, 95%, 97% or 98% identical to thefingerprint of NUN 09050. In a preferred embodiment amplified fragmentlength polymorphism (AFLP) markers are used for DNA fingerprinting (Voset al. 1995, Nucleic Acid Research 23: 4407-4414). A closely relatedplant may have a Jaccard's Similarity index of at least about 0.8,preferably at least about 0.9, 0.95, 0.96, 0.97, 0.98 or more (vanEeuwijk and Law (2004), Euphytica 137: 129-137). In one embodiment aclosely related plant of NUN 09050 has a Jaccard Similarity indez ofhigher than 0.96. See also the Guidelines on Essentially DerivedVarieties for lettuce published by the ISF(http://www.worldseed.org/isf/edv.html), where the technical protocolfor assessing the Jaccard index is outlined. The following 10 AFLPprimer combinations can be used: E33/M59, E35/M48, E35/M49, E35/M59,E35/M60, E38/M54, E44/M48, E44/M49, E45/M48, E45/M49.

By crossing and/or selfing also (one or more) single traits may beintroduced into NUN 09050 (e.g., using backcrossing breeding schemes),while retaining the remaining morphological and physiologicalcharacteristics of NUN 09050. For example, disease resistance genes maybe introduced, genes responsible for one or more quality traits (such ashead quality), yield, etc. Both single genes (dominant. semidominant orrecessive) and one or more QTLs (quantitative trait loci) may betransferred into NUN 09050 by breeding with NUN 09050.

Any pest or disease resistance genes may be introduced into NUN 09050,progeny thereof or into an EDV of NUN 09050. Resistance against one ormore of the following diseases is preferably introduced into plants ofthe invention: downy mildew, Sclerotinia rot, Botrytis, powdery mildew,anthracnose, bottom rot, corky root rot, lettuce mosaic virus, big vein,lettuce aphid, beet western yellows and aster yellows. Resistanceagainst one or more of the following pests is preferably present orintroduced into plants of the invention: Sclerotinia minor (leaf drop),Sclerotinia sclerotiorum (leaf drop), Rhizoctonia solani (bottom drop),Erysiphe cichoracearum (powdery mildew), Fusarium oxysporum f. sp.lactucae (Fusarium wilt) resistance. Other resistance genes, againstpathogenic viruses (e.g. Lettuce infectious yellows virus (LIYV),lettuce mosaic virus (LMV), Cucumber mosaic virus (CMV), Beet westernyellows virus (BWYV), Alfalfa mosaic virus (AMY)), fungi, bacteria orlettuce pests may also be introduced. In one embodiment resistanceagainst Nasonovia ribisnigri biotype Nr:0 and/or Nr:1 is introduced intoNUN 09050.

Transgene

Also provided is a method of producing a lettuce plant having a desiredtrait, wherein the method comprises transforming the lettuce plant ofthe invention with a transgene that confers the desired trait, whereinthe transformed plant retains essential all phenotypic and morphologicalcharacteristics of a NUN 09050 plant of the invention and contains thedesired trait. Thus, a transgenic lettuce plant is provided which isproduced by the method described above, wherein the plant comprises thedesired trait and essentially all of the physiological and morphologicalcharacteristics of a NUN 09050 plant.

Many useful traits that can be introduced into NUN 09050 by e.g.crossing NUN 09050 with a transgenic lettuce plant comprising a desiredtransgene, as well as by directly introducing a transgene into NUN 09050(or an EDV, or progeny of NUN 09050) by genetic transformationtechniques. Genetic transformation may, therefore, be used to insert aselected transgene into the lettuce plants of the invention or may,alternatively, be used for the preparation of transgenic lettuce plantswhich can then be used as a source of the transgene(s), which can beintroduced into NUN 09050 by e.g. backcrossing. Methods for thetransformation of plants, including lettuce, are well known to those ofskill in the art.

Any DNA sequences, whether from a different species or from the samespecies, which are inserted into the genome using transformation, arereferred to herein collectively as “transgenes”. A “transgene” alsoencompasses antisense, or sense and antisense sequences capable of genesilencing. Thus, the present invention also relates to transgenic NUN09050 plants. In some embodiments of the invention, a transgenic NUN09050 plants may contain at least one transgene but could contain atleast 1, 2, 3, 4, 5, 6, or more transgenes.

One embodiment of the invention is a process for producing progeny of aplant designated NUN 09050 further comprising a desired trait, saidprocess comprising transforming a NUN 09050 plant (or a cell or tissueof NUN 09050) with at least one transgene that confers a desired traitand/or crossing a plant designated NUN 09050 with a transgenic lettuceplant comprising a desired transgene so that the genetic material of theprogeny that results from the cross contains the transgene(s). The DNAof the transgene is operatively linked to a regulatory element active inplant cells (especially a promoter) and the transgene confers a traitselected from the group consisting of male sterility, male fertility,herbicide resistance, insect resistance, disease resistance, improvedyield, improved nutritional quality or improved biotic or abiotic (e.g.drought or salt) stress tolerance. Another embodiment is the productproduced by this process. In one embodiment the desired trait which isbased on a transgene may be one or more of herbicide resistance, insectresistance, disease resistance, stress tolerance, modified fatty acid ormodified carbohydrate metabolism. The specific transgene may be anyknown in the art or listed herein, including, a polynucleotide sequenceconferring resistance to imidazolinone, sulfonylurea, glyphosate,glufosinate, triazine, benzonitrile, cyclohexanedione, phenoxyproprionic acid and L-phosphinothricin or a polynucleotide conferringresistance to nematodes, downy mildew, Sclerotinia rot, Botrytis,powdery mildew, anthracnose, bottom rot, corky root rot, lettuce mosaicvirus, big vein, lettuce aphid, beet western yellows and aster yellows,Sclerotinia minor (leaf drop), Sclerotinia sclerotiorum (leaf drop),Rhizoctonia solani (bottom drop), Erysiphe cichoracearum (powderymildew), Fusarium oxysporum f. sp. Lactucae (fusarium wilt), lettuceinfectious yellows virus (LIYV), lettuce mosaic virus (LMV), Cucumbermosaic virus (CMV), Beet western yellows virus (BWYV), and Alfalfamosaic virus (AMV).

Numerous methods for plant transformation have been developed, includingbiological and physical plant transformation protocols (see, e.g.,Glick, B. R. and Thompson, J. E. Eds. (CRC Press, Inc., Boca Raton(1993): 67-88 and Armstrong, “The First Decade of Maize Transformation:A Review and Future Perspective” Maydica 1999 (44):101-109). Inaddition, expression vectors and in vitro culture methods for plant cellor tissue transformation and regeneration of plants are available. See,e.g., Glick, B. R. and Thompson, J. E. Eds. (CRC Press, Inc., Boca Raton(1993): 89-119).

A genetic trait which has been engineered into the genome of aparticular lettuce plant may then be moved into the genome of anotherlettuce plant (e.g. another variety) using traditional breedingtechniques that are well known in the plant breeding arts. For example,a backcrossing approach is commonly used to move a transgene from atransformed lettuce variety into an already developed lettuce variety,and the resulting backcross conversion plant will then comprise thetransgene(s).

Various genetic elements can be introduced into the plant genome usingtransformation. These elements include, but are not limited to genes,coding sequences, inducible-, constitutive-, and tissue specificpromoters, enhancing sequences, and signal and targeting sequences. Forexample, see the traits, genes, and transformation methods listed inU.S. Pat. No. 6,118,055.

Plant transformation involves the construction of an expression vectorwhich will function in plant cells. Such a vector comprises DNAcomprising a gene under control of, or operatively linked to, aregulatory element (for example a promoter). The expression vector maycontain one or more such operably linked gene/regulatory elementcombinations. The vector(s) may be in the form of a plasmid and can beused alone or in combination with other plasmids to provide transformedlettuce plants using transformation methods as described below toincorporate transgenes into the genetic material of the lettuceplant(s).

Although also marker-free transformation methods are known, expressionvectors generally include at least one genetic marker operably linked toa regulatory element (a promoter, for example) that allows transformedcells containing the marker to be either recovered by negativeselection, i.e., inhibiting growth of cells that do not contain theselectable marker gene, or by positive selection, i.e., screening forthe product encoded by the genetic marker. Thus, vectors used for thetransformation of lettuce cells are not limited so long as the vectorcan express an inserted DNA in the cells. For example, vectorscomprising promoters for constitutive gene expression in lettuce cells(e.g., cauliflower mosaic virus 35S promoter) and promoters inducible byexogenous stimuli can be used. Examples of suitable vectors include pBIbinary vector. The “lettuce cell” into which the vector is to beintroduced includes various forms of lettuce cells, such as for examplecultured cell suspensions, protoplasts, leaf sections, and callus.

Many commonly used selectable marker genes for plant transformation arewell known in the transformation arts, and include, for example, genesthat code for enzymes that metabolically detoxify a selective chemicalagent which may be an antibiotic or a herbicide, or genes that encode analtered target which is insensitive to the inhibitor. A few positiveselection methods are also known in the art.

One commonly used selectable marker gene for plant transformation is theneomycin phosphotransferase II (nptII) gene which, when under thecontrol of plant regulatory signals, confers resistance to kanamycin(see, e.g., Fraley et al., Proc. Natl. Acad. Sci. USA (1983), 80:4803-4807). Another commonly used selectable marker gene is thehygromycin phosphotransferase gene which confers resistance to theantibiotic hygromycin (see, e.g., Vanden Elzen et al., Plant Mol. Biol,(1985) δ: 299-302).

Additional selectable marker genes of bacterial origin that conferresistance to antibiotics include gentamycin acetyl transferase,streptomycin phosphotransferase and aminoglycoside-3′-adenyltransferase, the bleomycin resistance determinant. Other selectablemarker genes confer resistance to herbicides such as glyphosate,glufosinate or bromoxynil. Further selectable markers such as mousedihydrofolate reductase and plant acetolactate synthase are well knownin the art. Also known are marker genes for plant transformation whichrequire screening of presumptively transformed plant cells rather thandirect genetic selection of transformed cells for resistance to a toxicsubstance such as an antibiotic. These genes are particularly useful toquantify or visualize the spatial pattern of expression of a gene inspecific tissues and are frequently referred to as reporter genesbecause they can be fused to a gene or gene regulatory sequence for theinvestigation of gene expression. The use of such markers is alsoencompassed by the present invention.

The gene encoding Green Fluorescent Protein (GFP) can also be utilizedas a marker for gene expression in prokaryotic and eukaryotic cells(Chalfie et al., Science (1994) 263: 802-805). GFP and mutants of GFPmay be used as screenable markers.

Genes included in expression vectors must be driven by a nucleotidesequence comprising a regulatory element, for example, a promoter.Several types of promoters are well known in the transformation arts asare other regulatory elements that can be used alone or in combinationwith promoters.

As used herein, “promoter” includes reference to a region of DNAupstream from the start of transcription and involved in recognition andbinding of RNA polymerase and other proteins to initiate transcription.A “plant promoter” is a promoter capable of initiating transcription inplant cells. Exemplary promoters are well known in the art such asinducible promotors, constitutive promoters, tissue-specific promotorsor tissue-preferred promoters. One example is the cauliflower mosaicvirus 35S promoter.

Transport of a protein produced by transgenes to a subcellularcompartment such as the chloroplast, vacuole, peroxisome, glyoxysome,cell wall or mitochondrion or for secretion into the apoplast, isaccomplished by means of operably linking the nucleotide sequenceencoding a signal sequence to the 5′ and/or 3′ region of a gene encodingthe protein of interest. Targeting sequences at the 5′ and/or 3′ end ofthe structural gene may determine during protein synthesis andprocessing where the encoded protein is ultimately compartmentalized.

The presence of a signal sequence directs a polypeptide to either anintracellular organelle (e.g. chloroplasts) or subcellular compartmentor for secretion to the apoplast. Many signal sequences are known in theart (see, e.g., Becker et al., Plant Mol. Biol. (1992) 20: 49-60; orKnox, C., et al., Plant Mol. Biol. (1987) 9: 3-17).

By means of the present invention, plants can be genetically engineeredto express various phenotypes of horticultural interest. Through thetransformation of lettuce the expression of genes can be altered toenhance disease resistance, insect resistance, herbicide resistance,stress tolerance, horticultural quality, and other traits.Transformation can also be used to insert DNA sequences which control orhelp control male sterility or fertility restoration. DNA sequencesnative to lettuce as well as non-native DNA sequences can be transformedinto lettuce and used to alter levels of native or non-native proteins.Various promoters, targeting sequences, enhancing sequences, and otherDNA sequences can be inserted into the genome for the purpose ofaltering the expression of proteins. Reduction of the activity ofspecific genes (also known as gene silencing, or gene suppression) isdesirable for several aspects of genetic engineering in plants.

Many techniques for gene silencing are well known to one of skill in theart, including, but not limited to, knock-outs (such as by insertion ofa transposable element such as mu) or other genetic elements such as aFRT, Lox or other site specific integration site, antisense technology,co-suppression, RNA interference, virus-induced gene silencing,target-RNA-specific ribozymes, ribozymes, oligonucleotide mediatedtargeted modification, Zinc-finger targeted molecules (e.g., WO 01/52620or WO 03/048345); and other methods or combinations of the above methodsknown to those of skill in the art.

Likewise, by means of the present invention, other genes can beexpressed in transformed plants. More particularly, plants can begenetically engineered to express various phenotypes of interest.

Exemplary nucleic acids which may be introduced to the lettuce lines ofthis invention include, for example, DNA sequences or genes from anotherspecies, or even genes or sequences which originate from, or are presentin, the same species, but are incorporated into recipient cells bygenetic engineering methods rather than classical reproduction orbreeding techniques. However, the term “exogenous” is also intended torefer to genes that are not normally present in the cell beingtransformed, or perhaps simply not present in the form, structure, etc.,as found in the transforming DNA segment or gene, or genes which arenormally present and that one desires to express in a manner thatdiffers from the natural expression pattern, e.g., to over-express.Thus, the term “exogenous” gene or DNA is intended to refer to any geneor DNA segment that is introduced into a recipient cell, regardless ofwhether a similar gene may already be present in such a cell. The typeof DNA included in the exogenous DNA can include DNA which is alreadypresent in the plant cell, DNA from another plant, DNA from a differentorganism, or a DNA generated externally, such as a DNA sequencecontaining an antisense message of a gene, or a DNA sequence encoding asynthetic or modified version of a gene.

Non-limiting examples of particular genes and corresponding phenotypesone may choose to introduce into a lettuce plant include one or moregenes for insect tolerance, such as a Bacillus thuringiensis (B.t.)gene, disease tolerance such as genes for fungal or bacterial diseasecontrol, herbicide tolerance such as genes conferring glyphosatetolerance, and genes for quality improvements such as yield, nutritionalenhancements, environmental or stress tolerances, or any desirablechanges in plant physiology, growth, development, morphology or plantproduct(s). A more detailed list of genes implicated in this regardincludes, but is not limited to, genes that confer resistance to pestsor disease and that encode:

-   -   plant disease resistance genes (plant defenses are often        activated by specific interaction between the product of a        disease resistance gene (R) in the plant and the product of a        corresponding avirulence (Avr) gene in the pathogen,    -   genes conferring resistance to a pest, such as a nematode (see,        e.g., WO 96/30517; WO 93/19181; WO 03/033651) or insects (e.g.,        insect-specific hormones or pheromones, or insect-specific        peptides or neuropeptides which, upon expression, disrupts the        physiology of the affected pest),    -   an enzyme involved in the modification, including the        post-translational modification, of a biologically active        molecule; for example, a glycolytic enzyme, a proteolytic        enzyme, a lipolytic enzyme, a nuclease, a cyclase, a        transaminase, an esterase, a hydrolase, a phosphatase, a kinase,        a phosphorylase, a polymerase, an elastase, a chitinase, and a        glucanase, whether natural or synthetic (see, e.g., WO        93/02197),    -   a viral-invasive protein or a complex toxin derived therefrom,        e.g., the accumulation of viral coat proteins in transformed        plant cells imparts resistance to viral infection and/or disease        development effected by the virus from which the coat protein        gene is derived, as well as by related viruses (see, e.g.,        Beachy et al., Ann. Rev. Phytopathol. (1990) 28: 451-474). Coat        protein-mediated resistance has been conferred upon transformed        plants against alfalfa mosaic virus, cucumber mosaic virus, and        tobacco mosaic virus),    -   a virus-specific antibody (see, e.g., Tavladoraki et al.,        Nature (1993) 366: 469-472), a developmental-arrestive protein        produced in nature by a pathogen or a parasite. Thus, fungal        endo-α-1,4-D-polygalacturonases facilitate fungal colonization        and plant nutrient release by solubilizing plant cell wall        homo-α-1,4-D-galacturonase (see, e.g., Lamb et al.,        Bio/Technology (1992) 10: 1436-1445),    -   genes involved in the Systemic Acquired Resistance (SAR)        Response and/or the pathogenesis-related genes or antifungal        genes (see, e.g., U.S. Pat. No. 6,875,907),    -   cystatin and cysteine proteinase inhibitors (see, e.g., U.S.        Pat. No. 7,205,453),    -   defensin genes (see, e.g., WO 03/000863 and U.S. Pat. No.        6,911,577),    -   genes conferring virus resistance, e.g. coat protein genes        conferring resistance against Lettuce big vein disease (Kawazu        et al, 2010, Transgenic Research Vol: 19(2): 211-220).

Also included, but not limited to, are genes that confer resistance toan herbicide, e.g.:

-   -   a herbicide that inhibits the growing point or meristem, such as        an imidazolinone or a sulfonylurea. Exemplary genes in this        category code for mutant ALS and AHAS enzyme (see, e.g., Lee et        al., EMBO J. (1988). 7: 1241-1248),    -   Glyphosate (resistance, e.g., conferred by mutant        5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) and aroA        genes, respectively, see, e.g. U.S. Pat. No. 5,776,760, U.S.        Pat. No. 5,463,175, U.S. Ser. No. 10/427,692 or U.S. Pat. No.        4,769,061) and other phosphono compounds such as glufosinate,        and pyridinoxy or phenoxy proprionic acids and cyclohexanediones        (see, e.g., U.S. Pat. No. 5,627,061, U.S. Pat. No. 6,566,587;        U.S. Pat. No. 6,338,961; U.S. Pat. No. 6,248,876; U.S. Pat. No.        6,040,497; U.S. Pat. No. 5,804,425; U.S. Pat. No. 5,633,435; EP        1 173 580; EP 1 173 581; and EP 1 173 582). EP 0 333 033, and        U.S. Pat. No. 4,975,374 disclose nucleotide sequences of        glutamine synthetase genes which confer resistance to herbicides        such as L-phosphinothricin. The nucleotide sequence of a PAT        gene is provided in EP 0 242 246. DeGreef et al.        (Bio/Technology (1989) 7: 61-64) describe the production of        transgenic plants that express chimeric bar genes coding for        phosphinothricin acetyl transferase activity.    -   a herbicide that inhibits HPPD (hydroxyphenylpyruvate        dioxygenase, e.g., WO 2009/144079),

a herbicide that inhibits photosynthesis, protoporphyrinogen oxidase(protox) enzyme serves as the target for a variety of herbicidalcompounds (see, e.g., U.S. Pat. No. 6,288,306; U.S. Pat. No. 6,282,837;U.S. Pat. No. 5,767,373; and WO 01/12825).

Also included, but not limited to, are genes that confer or contributeto a value-added trait, such as:

-   -   modified fatty acid metabolism,

decreased phytate,

modified carbohydrate composition,

altered antioxidant content or composition,

Also included, but not limited to, are genes that control male sterility(see, e.g., U.S. Pat. No. 4,654,465).

Also included, but not limited to, are genes that create a site for sitespecific DNA integration (see, e.g., WO 99/25821), genes that affectabiotic stress resistance, and other genes and transcription factorsthat affect plant growth and other traits such as yield, flowering,plant growth, and/or plant structure, can be introduced or introgressedinto plants.

Transgenic lettuce can also be used as protein-factory, e.g. for theproduction of edible vaccines.

A vector can be introduced into lettuce cells by known methods, such asthe polyethylene glycol method, polycation method, electroporation,Agrobacterium-mediated transfer, microprojectile bombardment and directDNA uptake by protoplasts.

To effect transformation by electroporation, one may employ eitherfriable tissues, such as a suspension culture of cells or embryogeniccallus or alternatively one may transform immature embryos or otherorganized tissue directly. In this technique, one would partiallydegrade the cell walls of the chosen cells by exposing them topectin-degrading enzymes (pectolyases) or mechanically wound tissues ina controlled manner. An example of electroporation of lettuceprotoplasts is presented in Chupeau et al. (1989; Bio/Technology 7:503-508).

Microprojectile bombardment techniques are widely applicable, and may beused to transform virtually any plant species. Examples involvingmicroprojectile bombardment transformation with lettuce can be found in,e.g., Elliott et al. Phys. Rev. Lett. (2004) 92: 095501.

When using microprojectile bombardment, particles are coated withnucleic acids and delivered into cells by a propelling force. Exemplaryparticles include those comprised of tungsten, platinum, and preferably,gold. For the bombardment, cells in suspension are concentrated onfilters or solid culture medium. Alternatively, immature embryos orother target cells may be arranged on solid culture medium. The cells tobe bombarded are positioned at an appropriate distance below themacroprojectile stopping plate.

An illustrative embodiment of a method for delivering DNA into plantcells by acceleration is the Biolistics Particle Delivery System, whichcan be used to propel particles coated with DNA or cells through ascreen, such as a stainless steel or Nytex screen, onto a surfacecovered with target lettuce cells. The screen disperses the particles sothat they are not delivered to the recipient cells in large aggregates.It is believed that a screen intervening between the projectileapparatus and the cells to be bombarded reduces the size of projectilesaggregate and may contribute to a higher frequency of transformation byreducing the damage inflicted on the recipient cells by projectiles thatare too large.

Agrobacterium-mediated transfer is another widely applicable system forintroducing transgenes into plant cells. Modern Agrobacteriumtransformation vectors are capable of replication in E. coli as wellknown in the art. For example, U.S. Pat. No. 5,349,124 describes amethod of transforming lettuce plant cells using Agrobacterium-mediatedtransformation. By inserting a chimeric gene having a DNA codingsequence encoding for the full-length B.t. toxin protein that expressesa protein toxic toward Lepidopteran larvae, this methodology resulted inlettuce having resistance against such insects.

Transformation of plant protoplasts also can be achieved using methodsbased on calcium phosphate precipitation, polyethylene glycol treatment,electroporation, and combinations of these treatments (see, e.g.,Neuhaus and Spangenberg, Physiologia Plantarum (1990) 79 (1): 213-217).

The foregoing methods for transformation would typically be used forproducing a transgenic plant. The transgenic plant could then be crossedwith another (non-transgenic or transgenic) plant in order to produce anew transgenic plant. Alternatively, a genetic trait that has beenengineered into a particular lettuce plant using the foregoingtransformation techniques could be moved into another plant usingtraditional breeding techniques that are well known in the plantbreeding arts.

Products and Package

Also provided are plant parts derived from variety NUN 09050, or from avegetatively propagated plant of NUN 09050, being selected from thegroup consisting of: harvested (mature or immature) leaves or partsthereof, pollen, ovules, cells, heads, cotyledons, seeds or partsthereof, stalks or parts thereof, roots or parts thereof, cuttings, orparts thereof, flowers, florets, or flower buds.

In one embodiment, the invention provides for extracts of a plantdescribed herein and compositions comprising or consisting of suchextracts. In a preferred embodiment, the extract consists of orcomprises tissue of a plant described herein or is obtained from suchtissue. For example sesquiterpene-lactones such as lactucin, lactucid orlactucopicrin; or triterpenes such as amyrin, lactucerol or taraxasterolcyanine may be an extract obtained from leaf tissue and used to make ahealth-beneficial composition (e.g., a pharmaceutical or nutraceuticalcomposition).

The invention also provides for a food or feed product comprising orconsisting of a plant part described herein and/or an extract from aplant part described herein. The food or feed product may be fresh orprocessed, e.g., canned, steamed, boiled, fried, blanched and/or frozenetc.

A lettuce plant designated NUN 09050, a progeny thereof, a derivedvariety thereof (such as EDV), and parts of the afore-mentionedplants/varieties can be suitably packed for, e.g., transport, and/orsold fresh. Such parts encompass any cells, tissues and organsobtainable from the seedlings or plants, such as but not limited to:heads, cuttings, pollen, leaves, parts of leaves, and the like. Headsand leaves may be harvested immature, as baby-leaf, or mature. A plant,plants or parts thereof may be packed in a container (e.g., bags,cartons, cans, etc.) alone or together with other plants or materials.Parts can be stored and/or processed further. Encompassed are thereforealso food or feed products comprising one or more of such parts, suchleaves or parts thereof obtainable from NUN 09050 plant, a progenythereof a derived variety thereof (such as EDV), and parts of theafore-mentioned plants/varieties.

For example, containers such as cans, boxes, crates, bags, cartons,Modified Atmosphere Packagings, films (e.g. biodegradable films), etc.comprising plant parts of plants (fresh and/or processed) designated NUN09050 are also provided herein.

Color

The color of a plant in accordance with the present invention can bedetermined by comparing the color of, e.g., fourth or mature leaves withfourth or mature leaves of a MULTY plant grown under the sameconditions. The skilled person can, e.g., use a RHS colour chart: 2007as described herein.

DEPOSIT INFORMATION

A total of 2500 seeds of the variety NUN 09050 (also called “MULTIGREEN50”) are deposited by Nunhems B.V. on ______, at the American TypeCulture Collection (ATCC), University Boulevard, Manassas, Va.20110-2209 USA and/or at the NCIMB Ltd., Ferguson Building, CraibstoneEstate, Bucksburn, Aberdeen AB21 9YA, United Kingdom (NCIMB). Thedeposit has been assigned Accession Number PTA ______or NCIMB ______. Adeposit of NUN 09050 and of the parent lines is also maintained atNunhems B.V. Access to the deposit will be available during the pendencyof this application to persons determined by the Director of the U.S.Patent Office to be entitled thereto upon request. Subject to 37 C.F.R.§1.808(b), all restrictions imposed by the depositor on the availabilityto the public of the deposited material will be irrevocably removed uponthe granting of the patent. The deposit will be maintained for a periodof 30 years, or 5 years after the most recent request or for theenforceable life of the patent whichever is longer, and will be replacedif it ever becomes nonviable during that period. Applicant does notwaive any rights granted under this patent on this application or underthe Plant Variety Protection Act (7 USC 2321 et seq.).

Various modifications and variations of the described products andmethods of the invention will be apparent to those skilled in the artwithout departing from the scope and spirit of the invention. Althoughthe invention has been described in connection with specific preferredembodiments, it should be understood that the invention as claimedshould not be unduly limited to such specific embodiments. Indeed,various modifications of the described modes for carrying out theinvention which are obvious to those skilled in plant breeding,chemistry, biology or related fields are intended to be within the scopeof the following claims.

EXAMPLES Development of NUN 09050

The variety NUN 09050 was developed from an initial cross between alettuce variety and a breeding line. The female and male parents werecrossed to produce hybrid (F₁) seeds. After the cross, F₁ plants wereself pollinated. From the second to the fifth generation pedigreeselection was performed. From the sixth to the ninth generation lineselection was performed.

Variety NUN 09050 has been observed for at least three generations indifferent trials on different locations and during seed increase and isuniform and stable.

The variety is, therefore, uniform and genetically stable. This has beenestablished through evaluation of horticultural characteristics.Independent seed production events resulted in no observable deviationin genetic stability.

The seeds of NUN 09050 can be grown to produce plants and parts thereof(e.g. heads or leaves). The variety NUN 09050 can be propagated by seedsor vegetative propagation.

MULTIGREEN 50 characteristics were compared with those of MULTY andGRAND RAPIDS according to standards of the U.S. Department ofAgriculture, Agricultural Marketing Service, Science and Technology,Plant Variety Protection Office, Beltsville, Md. 20705. The trials werecarried out by Nunhems USA Inc. in Salinas (USA) (Planting date: Jun.26, 2011, duration 42 days, respectively 45 days).

Characteristics of NUN 09050

Table 1 shows the USDA descriptors of NUN 09050 (MULTIGREEN 50; thisapplication), MULTY and GRAND RAPIDS. 20 plants or plant parts wererandomly selected from two replications of 100 plants each. These 20plants were used to measure characteristics. The values are mean values.Some of the most significant differences are highlighted in bold.

TABLE 1 USDA NUN GRAND number USDA descriptor 09050 MULTY RAPIDS 1 Planttype 1 1 1 1 = Cutting/Leaf 2 Seed Color 2 2 2 1 = white, 2 = black, 3 =brown Light dormacy 1 — 2 1 = light required; 2 = light not requiredHead dormacy 2 — 1 1 = suseptible; 2 = not suseptible 3 Cotyledon tofourth leaf stage Shape of Cotyledons 2 3 2 1 = broad, 2 = intermediate,3 = spatulate Shape of fourth leaf 6 6 3 3 = oval, 6 = pinnately lobedLength/width index of fourth leaf (L/W*10) 90.6 89.2 116.2 Apical margin5 2 8 1 = entire, 2 = crenate/gnawed, 3 = finely dentate, 4 = moderatelydentate, 5 = coarsely dentate, 6 = incised, 7 = lobed, 8 = other(undulate) Basal margin 7 7 5 1 = entire, 2 = crenate/gnawed, 3 = finelydentate, 4 = moderately dentate, 5 = coarsely dentate, 6 = incised, 7 =lobed, 8 = other (undulate) Undulation 3 3 3 1 = flat, 2 = slight, 3 =medium, 4 = marked Green color 3 2 1 1 = yellow green, 2 = light green,3 = medium green, 4 = dark green Anthocyanin: Distribution 1 1 1 1 =absent, 2 = margin only, 3 = spotted, 4 = throughout Concentration — — —1 = light, 2 = moderate, 3 = intense Rolling 1 1 1 1 = absent, 2 =present Cupping 1 1 1 1 = uncupped, 3 = markedly Reflexing 1 1 2 2 =apical margin, 3 = lateral margins 4 Mature leaves (harvest mature outerleaves): Margin: Incision depth 2 3 2 1 = absent/shallow (Dark GreenBoston), 2 = moderate (Vanguard), 3 = deep (Great Lakes 659) Indentation2 3 2 1 = entire, 2 = shallowly dentate (Great Lake 65), 3 = deeplydentate (Great Lake 659) Undulations of the apical margin 3 3 3 1 =absent/slight (Dark Green Boston), 2 = moderate (Vanguard), 3 = strong(Great Lakes 659) Green color 4 4 1 1 = very light green, 2 = lightgreen, 3 = medium green, 4 = dark green, 6 = other Anthocyanin:Distribution 1 1 1 1 = absent, 3 = spotted (California Cream Butter), 4= throughout (Prize Head) Concentration — — — 1 = light, 2 = moderate, 3= intense Size 1 1 3 1 = small, 2 = medium, 3 = large Glossiness 2 3 2 1= dull, 2 = moderate, 3 = glossy Blistering 1 1 3 1 = absent/slight, 2 =moderate, 3 = strong Leaf thickness 2 2 3 1 = thin, 2 = intermediate, 3= thick Trichomes 1 1 1 1 = absent, 2 = present 5 Plant Spread of frameleaves 28.1 cm 21.2 cm 38.0 cm Head diameter (market trimmed with single— cm — cm — cm cap leaf) Head shape 5 5 5 1 = flattened, 4 = elongate, 5= non-heading Head size class — — — 1 = small, 2 = medium, 3 = largeHead per carton — — — Head weight 267.0 g 137.5 g 397.0 g Head firmness1 1 1 1 = loose, 3 = Firm, 4 = very firm 6 Butt Shape — — — 1 = slightlyconcave, 2 = flat, 3 = rounded Midrib — — — 3 = prominently raised 7Core Diameter at base of head 19.8 mm 14.0 mm 31.4 mm Ratio of spreadframe leaves/core diameter 1.4 1.6 3.4 Core height from base of head toapex 39.2 mm 19.2 mm 54.0 mm 8 Bolting (not measured (—)) Number of daysfrom first water date to seed — — — stalk emergence (note: First waterdate is the date seed first receives adequate moisture to germinate.This often equals planting date). Bolting class — — — 1 = very low, 3 =medium, 4 = rapid, 5 = very rapid Height of mature seed stalk — — —Spread of bolter plant (at widest point) — — — Color — — — 1 = lightgreen, 2 = medium green 9 Maturity (earliness of harvest-mature head 42days 45 days 45 days formation) 10 Adaptation: — — — Season Fall (2), 0= not tested, 1 = not adapted, 2 = adapted Winter (0), Spring (2),Summer (0) Greenhouse 2 — — 0 = not tested, 1 = not adapted, 2 = adaptedSoil type 3 — — 1 = mineral, 2 = organic, 3 = both 11 Viral Diseases: —— — 12 Fungal/bacterial diseases: — — — Downy Mildew — — — (Races Bl: 1,5-13, 15, 17, 18, 27) 1 = immune, 3 = resistant, 5 = moderatelyresistant, 7 = susceptible, 9 = highly susceptible 13 Insects: 14Physiological stresses: — — — 1 = immune, 3 = resistant, 5 = moderatelyresistant, 7 = susceptible, 9 = highly susceptible Tipburn — — — 1 =immune; 5 = moderately resistant, 7 = susceptible, 9 = highlysusceptible Heat — — — 1 = immune; 5 = moderately resistant, 7 =susceptible, 9 = highly susceptible Brown rib — — — (Rib Discoloration,Rib Blight) Pink Rib — — — 1 = immune; 5 = moderately resistant, 7 =susceptible, 9 = highly susceptible Russet Spotting — — — 1 = immune; 5= moderately resistant, 7 = susceptible, 9 = highly susceptible — = notmeasured

These are typical values. Values may vary due to environment. Othervalues that are substantially equivalent are also within the scope ofthe invention.

Without limiting the scope of the present invention, picture 1 shows thedifferent color of fourth leaves of NUN 09050, MULTY and Grand RAPIDS aswell as the different shapes of leaves of said varieties.

1. A lettuce plant, designated NUN 09050, or part thereof, arepresentative sample of seeds of which having been deposited underAccession Number NCIMB ______ or PTA ______.
 2. A seed designated NUN09050, a representative sample of seeds having been deposited underAccession Number NCIMB ______ or PTA ______.
 3. A plant, or a partthereof, produced by growing the seed of claim
 2. 4. A part of the plantof claim 1, wherein the part is a head or a leaf or a part thereof.
 5. Acell culture or tissue culture of the lettuce plant of claim
 1. 6. Alettuce plant which is regenerated from the part of the plant of claim 1and comprises essentially all physiological and morphologicalcharacteristics of NUN
 09050. 7. A lettuce plant which is regeneratedfrom the cell culture or tissue culture of claim 5 and comprisesessentially all physiological and morphological characteristics of NUN09050.
 8. A progeny plant of lettuce variety designated NUN 09050obtained by further breeding with said variety, wherein said progenyplant has essentially all physiological and morphologicalcharacteristics of the variety designated NUN 09050 when grown under thesame environmental conditions.
 9. An Essentially Derived Variety of NUN09050 having at least one, two or three physiological and/ormorphological characteristics which are different from those of NUN09050 and which otherwise has essentially all physiological andmorphological characteristics of a lettuce plant designated NUN 09050.10. An Essentially Derived Variety of NUN 09050 according to claim 9having one or two physiological and/or morphological characteristicswhich are different from those of NUN 09050 and which otherwise hasessentially all physiological and morphological characteristics of alettuce plant designated NUN 09050 obtainable by selecting a natural orinduced mutant, or a somaclonal variant from a population of plantsdesignated NUN
 09050. 11. A package comprising a head, leaves or partsthereof of a plant of claim
 1. 12. A package comprising the seed ofclaim
 2. 13. A seed pellet comprising the seed of claim
 2. 14. A methodof producing a lettuce plant, comprising crossing the plant of claim 1with a second lettuce plant one or more times and/or selfing the plantof claim 1 one or more times, and selecting progeny from said crossingand/or selfing.
 15. A method of producing a lettuce plant derived fromNUN 09050 comprising the steps of: (a) preparing a progeny plant derivedfrom NUN 09050 by crossing the plant of claim 1 with itself or with asecond lettuce plant; (b) crossing the progeny plant with itself or asecond lettuce plant to produce a seed of a progeny plant of asubsequent generation; (c) growing a progeny plant of a subsequentgeneration from said seed and crossing the progeny plant of a subsequentgeneration with itself or a second lettuce plant; and (d) repeating stepb) and/or c) for at least 1 more generation to produce a lettuce plantderived from NUN
 09050. 16. A method of producing a hybrid lettuce seed,comprising crossing a first parent lettuce plant with a second parentlettuce plant and harvesting the resultant hybrid lettuce seed, whereinsaid first parent lettuce plant or said second parent lettuce plant is alettuce plant of claim
 1. 17. A hybrid lettuce seed produced by crossinga first parent lettuce plant with a second parent lettuce plant andharvesting the resultant hybrid lettuce seed, wherein said first parentlettuce plant or said second parent lettuce plant is a lettuce plant ofclaim
 1. 18. A method of introducing a single locus conversion into alettuce plant designated NUN 09050 comprising: (a) crossing a plantdesignated NUN 09050 with a second plant comprising a desired singlelocus to produce a F₁ progeny plant; (b) optionally selfing said F₁progeny plant to produce a F₂ progeny plant having said single locus;(c) crossing said F₁ or F₂ progeny plant of step (a) or step (b),respectively, with a plant of NUN 09050, representative seed of whichhaving been deposited under Accession Number NCIMB ______ or PTA ______,to produce backcross progeny plants; (d) selecting backcross progenyplants that have the single locus and otherwise essentially allphysiological and morphological characteristics of a plant designatedNUN 09050; and (e) repeating steps (c) and (d) one or more times insuccession to produce selected second or higher backcross progeny plantsthat comprise the single locus and otherwise comprise essentially allphysiological and morphological characteristics of a plant designatedNUN
 09050. 19. The method of claim 18, wherein the single locus confersa trait, wherein the trait is pest resistance or disease resistance. 20.The method of claim 18, wherein the single locus confers at least onetrait, wherein said trait is at least one trait selected from the groupconsisting of resistance against Nasonovia ribisnigri Nr: 1 or Nr: 0,downy mildew, Sclerotinia rot, Botrytis, powdery mildew, anthracnose,bottom rot, corky root rot, lettuce mosaic virus, big vein, lettuceaphid, beet western yellows and aster yellows, Sclerotinia minor (leafdrop), Sclerotinia sclerotiorum (leaf drop), Rhizoctonia solani (bottomdrop), Erysiphe cichoracearum (powdery mildew), Fusarium oxysporum f.sp. Lactucae (fusarium wilt), lettuce infectious yellows virus (LIYV),lettuce mosaic virus (LMV), Cucumber mosaic virus (CMV), Beet westernyellows virus (BWYV), Alfalfa mosaic virus (AMV), nematodes andherbicides.
 21. A plant of claim 1 wherein said plant has at least theessential physiological and morphological characteristics of the varietydesignated NUN 09050 and further comprises at least one transgene thatconfers a desired trait.